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Metabolic Coupling Determines the Activity: Comparison of 11β-Hydroxysteroid Dehydrogenase 1 and Its Coupling between Liver Parenchymal Cells and Testicular Leydig Cells.

Li X, Hu G, Li X, Wang YY, Hu YY, Zhou H, Latif SA, Morris DJ, Chu Y, Zheng Z, Ge RS - PLoS ONE (2015)

Bottom Line: S3483, a G6P transporter inhibitor, reversed the G6P-mediated increases of 11β-HSD1 reductase activity.The depletion of Leydig cells eliminated Hsd11b1 (encoding 11β-HSD1) expression but did not affect the expression of H6pd (encoding H6PDH) and Slc37a4 (encoding G6P transporter).In conclusion, the availability of H6PDH determines the different direction of 11β-HSD1 in liver and Leydig cells.

View Article: PubMed Central - PubMed

Affiliation: The Second Affiliated Hospital & Yuying Children's Hospital, Wenzhou Medical University, Wenzhou, ZJ 325000, PR China.

ABSTRACT

Background: 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1) interconverts active 11β-hydroxyl glucocorticoids and inactive 11keto forms. However, its directionality is determined by availability of NADP+/NADPH. In liver cells, 11β-HSD1 behaves as a primary reductase, while in Leydig cells it acts as a primary oxidase. However, the exact mechanism is not clear. The direction of 11β-HSD1 has been proposed to be regulated by hexose-6-phosphate dehydrogenase (H6PDH), which catalyzes glucose-6-phosphate (G6P) to generate NADPH that drives 11β-HSD1 towards reduction.

Methodology: To examine the coupling between 11β-HSD1 and H6PDH, we added G6P to rat and human liver and testis or Leydig cell microsomes, and 11β-HSD1 activity was measured by radiometry.

Results and conclusions: G6P stimulated 11β-HSD1 reductase activity in rat (3 fold) or human liver (1.5 fold), but not at all in testis. S3483, a G6P transporter inhibitor, reversed the G6P-mediated increases of 11β-HSD1 reductase activity. We compared the extent to which 11β-HSD1 in rat Leydig and liver cells might be coupled to H6PDH. In order to clarify the location of H6PDH within the testis, we used the Leydig cell toxicant ethane dimethanesulfonate (EDS) to selectively deplete Leydig cells. The depletion of Leydig cells eliminated Hsd11b1 (encoding 11β-HSD1) expression but did not affect the expression of H6pd (encoding H6PDH) and Slc37a4 (encoding G6P transporter). H6pd mRNA level and H6PDH activity were barely detectable in purified rat Leydig cells. In conclusion, the availability of H6PDH determines the different direction of 11β-HSD1 in liver and Leydig cells.

No MeSH data available.


Related in: MedlinePlus

Effects of G6P and NADPH on rat liver and Leydig cell microsomal 11β-HSD1 reductase activities.11β-HSD1 reductase activity was measured using the microsomes of rat liver cells (Panel A) or Leydig cells (Panel B). Mean ± SEM, n = 5. Identical letters designate that there were no significant differences between two groups at P < 0.05.
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pone.0141767.g002: Effects of G6P and NADPH on rat liver and Leydig cell microsomal 11β-HSD1 reductase activities.11β-HSD1 reductase activity was measured using the microsomes of rat liver cells (Panel A) or Leydig cells (Panel B). Mean ± SEM, n = 5. Identical letters designate that there were no significant differences between two groups at P < 0.05.

Mentions: Rat liver microsome 11β-HSD1 reductase activity was measured in the presence of 0 to 400 μM G6P. G6P concentration-dependently increased liver microsome 11β-HSD1 reductase activity with 100 μM as a saturated concentration threshold (S2 Fig). However, addition of 0 to 400 μM G6P to rat Leydig cell microsomes had almost no effect on the velocity of the reaction (S2 Fig). The reductase activity of 11β-HSD1 in rat liver microsomes was stimulated significantly by 200 μM G6P after 10 min incubation. The addition of 1 μM S3483 (Fig 2A) reversed the stimulation by G6P. S3483 alone did not affect rat liver 11β-HSD1 oxidase and reductase activity when it was used up to 100 μM (S3 Fig). In contrast, G6P had no effects of 11β-HSD1 in rat Leydig cell microsomes (Fig 2B). We measured the NADP+ and NADPH levels in intact liver and Leydig cells, and found that both cells had the similar levels of NADP+ and NADPH (S1 Table). We further added 0.2 mM NADPH to rat liver and Leydig cell microsomes. NADPH increased 11β-HSD1 reductase activity by 2 folds, while NADPH stimulated 11β-HSD1 reductase activity in rat Leydig cell microsomes by 4 folds (Fig 2). We infer that rat Leydig cell 11β-HSD1 reductase is more dependent on NADPH than G6P.


Metabolic Coupling Determines the Activity: Comparison of 11β-Hydroxysteroid Dehydrogenase 1 and Its Coupling between Liver Parenchymal Cells and Testicular Leydig Cells.

Li X, Hu G, Li X, Wang YY, Hu YY, Zhou H, Latif SA, Morris DJ, Chu Y, Zheng Z, Ge RS - PLoS ONE (2015)

Effects of G6P and NADPH on rat liver and Leydig cell microsomal 11β-HSD1 reductase activities.11β-HSD1 reductase activity was measured using the microsomes of rat liver cells (Panel A) or Leydig cells (Panel B). Mean ± SEM, n = 5. Identical letters designate that there were no significant differences between two groups at P < 0.05.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4631333&req=5

pone.0141767.g002: Effects of G6P and NADPH on rat liver and Leydig cell microsomal 11β-HSD1 reductase activities.11β-HSD1 reductase activity was measured using the microsomes of rat liver cells (Panel A) or Leydig cells (Panel B). Mean ± SEM, n = 5. Identical letters designate that there were no significant differences between two groups at P < 0.05.
Mentions: Rat liver microsome 11β-HSD1 reductase activity was measured in the presence of 0 to 400 μM G6P. G6P concentration-dependently increased liver microsome 11β-HSD1 reductase activity with 100 μM as a saturated concentration threshold (S2 Fig). However, addition of 0 to 400 μM G6P to rat Leydig cell microsomes had almost no effect on the velocity of the reaction (S2 Fig). The reductase activity of 11β-HSD1 in rat liver microsomes was stimulated significantly by 200 μM G6P after 10 min incubation. The addition of 1 μM S3483 (Fig 2A) reversed the stimulation by G6P. S3483 alone did not affect rat liver 11β-HSD1 oxidase and reductase activity when it was used up to 100 μM (S3 Fig). In contrast, G6P had no effects of 11β-HSD1 in rat Leydig cell microsomes (Fig 2B). We measured the NADP+ and NADPH levels in intact liver and Leydig cells, and found that both cells had the similar levels of NADP+ and NADPH (S1 Table). We further added 0.2 mM NADPH to rat liver and Leydig cell microsomes. NADPH increased 11β-HSD1 reductase activity by 2 folds, while NADPH stimulated 11β-HSD1 reductase activity in rat Leydig cell microsomes by 4 folds (Fig 2). We infer that rat Leydig cell 11β-HSD1 reductase is more dependent on NADPH than G6P.

Bottom Line: S3483, a G6P transporter inhibitor, reversed the G6P-mediated increases of 11β-HSD1 reductase activity.The depletion of Leydig cells eliminated Hsd11b1 (encoding 11β-HSD1) expression but did not affect the expression of H6pd (encoding H6PDH) and Slc37a4 (encoding G6P transporter).In conclusion, the availability of H6PDH determines the different direction of 11β-HSD1 in liver and Leydig cells.

View Article: PubMed Central - PubMed

Affiliation: The Second Affiliated Hospital & Yuying Children's Hospital, Wenzhou Medical University, Wenzhou, ZJ 325000, PR China.

ABSTRACT

Background: 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1) interconverts active 11β-hydroxyl glucocorticoids and inactive 11keto forms. However, its directionality is determined by availability of NADP+/NADPH. In liver cells, 11β-HSD1 behaves as a primary reductase, while in Leydig cells it acts as a primary oxidase. However, the exact mechanism is not clear. The direction of 11β-HSD1 has been proposed to be regulated by hexose-6-phosphate dehydrogenase (H6PDH), which catalyzes glucose-6-phosphate (G6P) to generate NADPH that drives 11β-HSD1 towards reduction.

Methodology: To examine the coupling between 11β-HSD1 and H6PDH, we added G6P to rat and human liver and testis or Leydig cell microsomes, and 11β-HSD1 activity was measured by radiometry.

Results and conclusions: G6P stimulated 11β-HSD1 reductase activity in rat (3 fold) or human liver (1.5 fold), but not at all in testis. S3483, a G6P transporter inhibitor, reversed the G6P-mediated increases of 11β-HSD1 reductase activity. We compared the extent to which 11β-HSD1 in rat Leydig and liver cells might be coupled to H6PDH. In order to clarify the location of H6PDH within the testis, we used the Leydig cell toxicant ethane dimethanesulfonate (EDS) to selectively deplete Leydig cells. The depletion of Leydig cells eliminated Hsd11b1 (encoding 11β-HSD1) expression but did not affect the expression of H6pd (encoding H6PDH) and Slc37a4 (encoding G6P transporter). H6pd mRNA level and H6PDH activity were barely detectable in purified rat Leydig cells. In conclusion, the availability of H6PDH determines the different direction of 11β-HSD1 in liver and Leydig cells.

No MeSH data available.


Related in: MedlinePlus